False negative results from using common PCR reagents


The sensitivity of the PCR reaction makes it ideal for use when identifying potentially novel viral infections in human disease. Unfortunately, this same sensitivity also leaves this popular technique open to potential contamination with previously amplified PCR products, or "carry-over"contamination.

PCR product carry-over contamination can be prevented with uracil-DNA-glycosylase (UNG), and it is for this reason that it is commonly included in many commercial PCR master-mixes. While testing the sensitivity of PCR assays to detect murine DNA contamination in human tissue samples, we inadvertently discovered that the use of this common PCR reagent may lead to the production of false-negative PCR results.FindingsWe show here that contamination with minute quantities of UNG-digested PCR product or any negative control PCR reactions containing primer-dimers regardless of UNG presence can completely block amplification from as much as 60ng of legitimate target DNA.

Conclusions: These findings could potentially explain discrepant results from laboratories attempting to amplify MLV-related viruses including XMRV from human samples, as none of the published reports used internal-tube controls for amplification.

The potential for false negative results needs to be considered and carefully controlled in PCR experiments, especially when the target copy number may be low - just as the potential for false positive results already is.

Author: Dean BacichKathryn SobekJessica CummingsAllison AtwoodDenise O'Keefe
Credits/Source: BMC Research Notes 2011, 4:457



Published on: 2011-10-27



News Provider: 7thSpace Interactive / EUPB Press Office

Copyright by the authors listed above - made available via BioMedCentral (Open Access). Please make sure to read our disclaimer prior to contacting 7thSpace Interactive. To contact our editors, visit our online helpdesk. If you wish submit your own press release, click here.

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